Knockdown of phosphatase and tensin homolog (PTEN) inhibits fatty acid oxidation and reduces very low density lipoprotein assembly and secretion in calf hepatocytes

Abstract

Dairy cows with fatty liver exhibit hepatic lipid accumulation and disturbances in fatty acid oxidation and lipid transport. Phosphatase and tensin homolog (PTEN), a lipid phosphatase, regulates intrahepatic fatty acid oxidation and lipid transport in mice. Whether PTEN play a role in fatty acid oxidation and very low density lipoprotein (VLDL) assembly in calf hepatocytes are unknown. Hepatocytes isolated from 3 healthy female Holstein calves (1 d old, 30–40 kg) were infected with empty adenovirus with green fluorescent protein for 48 h (Ad-GFP group) or infected with PTEN knockdown adenovirus for 48 h (Ad-shPTEN group), or cultured in RPMI-1640 without Ad-shPTEN or Ad-GFP (control group). Compared with the Ad-GFP group, PTEN knockdown decreased mRNA and protein abundance and the activity of fatty acid oxidation-related molecules, including acyl-coA synthetase long-chain 1, carnitine palmitoyltransferase 1, carnitine palmitoyltransferase 2, and 3-hydroxy acyl-coA dehydrogenase. Furthermore, PTEN knockdown decreased mRNA and protein abundance of VLDL assembly-related molecules, including apolipoprotein B100, apolipoprotein E, microsomal triglyceride transfer protein, and low density lipoprotein receptor. Importantly, PTEN knockdown promoted triglyceride accumulation in hepatocytes and reduced the VLDL content in culture medium. A subsequent study was conducted on the following 4 groups: cells infected with Ad-GFP for 48 h and then treated with 2% BSA for another 24 h (Ad-GFP + BSA); cells infected with Ad-GFP for 48 h and then treated with 1.2 mM free fatty acids (FFA) and 2% BSA for another 24 h (Ad-GFP + 1.2 mM FFA); cells infected with Ad-shPTEN for 48 h and then treated with 2% BSA for another 24 h (Ad-shPTEN + BSA); cells infected with Ad-shPTEN for 48 h and then treated with 1.2 mM FFA and 2% BSA for another 24 h (Ad-shPTEN + 1.2 mM FFA). Compared with Ad-GFP + BSA, the abundances of PTEN and of fatty acid oxidation- and VLDL assembly-related proteins were lower in the Ad-GFP + 1.2 mM FFA group. Importantly, PTEN knockdown heightened the increase in triglyceride accumulation of hepatocytes and the decrease in VLDL content in culture medium induced by FFA. Overall, these in vitro data indicate that FFA inhibits PTEN expression, leading to triglyceride accumulation and the inhibition of VLDL assembly in calf hepatocytes. These findings suggest that PTEN may be a potential therapeutic target for FFA-induced hepatic steatosis in dairy cows.